Aromatic O-glycosylation

Carbohydrates carrying an aromatic aglycon are important natural products and thus key synthetic targets. However, due to the electron-withdrawing properties of aromatic rings, phenols are difficult to glycosylate. This review covers the most common carbohydrate donors used for aromatic O-glycosylation (anomeric acetates, halides, trichloroacetimidates and thioglycosides) as well as some less common donors. The scope of the review is to give practical examples of aromatic O-glycosylations and to offer guidelines for glycosylation of typical aromatic residues. Anomeric acetates or trichloroacetimidates, activated under acidic conditions, are preferred for electron rich aromatic aglycons, while glycosyl halides, activated using basic conditions, are preferred for electron deficient aromatic residues.     

Attenuation of pattern recognition receptor signaling is mediated by a MAP kinase kinase kinase

Pattern recognition receptors (PRRs) play a key role in plant and animal innate immunity. PRR binding of their cognate ligand triggers a signaling network and activates an immune response. Activation of PRR signaling must be controlled prior to ligand binding to prevent spurious signaling and immune activation. Flagellin perception in Arabidopsis through FLAGELLIN‐SENSITIVE 2 (FLS2) induces the activation of mitogen‐activated protein kinases (MAPKs) and immunity. However, the precise molecular mechanism that connects activated FLS2 to downstream MAPK cascades remains unknown. Here, we report the identification of a differentially phosphorylated MAP kinase kinase kinase that also interacts with FLS2. Using targeted proteomics and functional analysis, we show that MKKK7 negatively regulates flagellin‐triggered signaling and basal immunity and this requires phosphorylation of MKKK7 on specific serine residues. MKKK7 attenuates MPK6 activity and defense gene expression. Moreover, MKKK7 suppresses the reactive oxygen species burst downstream of FLS2, suggesting that MKKK7‐mediated attenuation of FLS2 signaling occurs through direct modulation of the FLS2 complex.     

Growth slow-down and growth arrest of human colon carcinoma cells HCT-8 in vitro after exposure to 5-fluoro-2'-deoxyuridine

Abstract. Cellular heterogeneity in drug response denotes a mixed response among individual cells in a drug treated population. Individual cell responses may be more complex than 'cell kill'and 'no response'. In this study we employed a colony formation assay and high-resolution image analysis to detect the various responses such as immediate and delayed cessation of growth, growth delay and growth slow-down, at the level of the individual colony. The evaluation was carried out using a human ileocaecal adenocarcinoma cell line (HCT-8) and the anti-tumour agent 5-fluoro-2'-deoxyuridine (FdUrd). In the presence of a drug concentration which, in standard monolayer assays, inhibits the growth to about 50% (IC₅₀) only about 20% of the colonies ceased to grow and the remaining colonies grew at a growth rate of about 70% of control. At an FdUrd concentration which, in standard monolayer assays, reduced growth by >90% (>IC₉₀), about 50% of the cells grew, with growth rates of about 30% of control. The slowing of growth, most prominent at lower drug concentrations, should be considered in determining mechanisms of drug action at the individual cell level. In clinical situations in which high drug doses are precluded by toxicity to normal tissues, growth slow-down may play a significant role in tumour response.     

Shape and volume of craniofacial cavities in intentional skull deformations

Intentional cranial deformations (ICD) have been observed worldwide but are especially prevalent in preColombian cultures. The purpose of this study was to assess the consequences of ICD on three cranial cavities (intracranial cavity, orbits, and maxillary sinuses) and on cranial vault thickness, in order to screen for morphological changes due to the external constraints exerted by the deformation device. We acquired CT‐scans for 39 deformed and 19 control skulls. We studied the thickness of the skull vault using qualitative and quantitative methods. We computed the volumes of the orbits, of the maxillary sinuses, and of the intracranial cavity using haptic‐aided semi‐automatic segmentation. We finally defined 3D distances and angles within orbits and maxillary sinuses based on 27 anatomical landmarks and measured these features on the 58 skulls. Our results show specific bone thickness patterns in some types of ICD, with localized thinning in regions subjected to increased pressure and thickening in other regions. Our findings confirm that volumes of the cranial cavities are not affected by ICDs but that the shapes of the orbits and of the maxillary sinuses are modified in circumferential deformations. We conclude that ICDs can modify the shape of the cranial cavities and the thickness of their walls but conserve their volumes. These results provide new insights into the morphological effects associated with ICDs and call for similar investigations in subjects with deformational plagiocephalies and craniosynostoses. Am J Phys Anthropol 151:110–119, 2013. © 2013 Wiley Periodicals, Inc.     

Determination of cortisol in human saliva using liquid chromatography-electrospray tandem mass spectrometry

The aim of this work was to develop a method for determination of cortisol in saliva by liquid chromatography-tandem mass spectrometry (LC-MS-MS). Saliva was sampled on Salivette tubes. These were centrifuged, deuterium-labeled cortisol was added as internal standard and the proteins precipitated by acetonitrile. The supernatant was evaporated, dissolved in methanol acidified with acetic acid and analyzed by LC-MS-MS. The with-in run precision, tested by pooling saliva samples from volunteers and then analyzing these in a single run, was found to be 7% at 0.7 microgram l(-1). The between-run precision was tested by analysis of the same samples at different days and found to be 11% at 2.5 microgram l(-1). The limit of quantification was 0.5 microgram l(-1). The method was applied for analysis of saliva samples from three volunteers during their last week before vacation and the first and second week on vacation. In addition, the method was compared to analysis by an immunological method. The values from the immunological method were 2.7 times higher than the LC-MS-MS results.     

Temperature-Sensitive Variants of NICOTIANA TABACUM Isolated from Somatic Cell Culture

Temperature-sensitive variants of Nicotiana tabacum were isolated from a liquid suspension culture of somatic cells by a negative selection procedure, using bromodeoxyuridine and light. A total of nine such variants have been recovered, with an estimated rate of 2 x 10^-7 per cell division. The appearance of the variants at the permissive temperature varied from nearly wild type, white and friable, to brown, compact and slow growing. Two of the variants adapted from growth on solid medium to growth in a liquid suspension culture; these were further characterized for chromosome number, growth rate, cell death rate at the restrictive temperature, growth on nutritionally modified media, and RNA and protein synthesis. The variants have been placed on regeneration media, and one of them has produced plantlets. Leaves from a plantlet have been placed on callus-inducing media, and the resulting callus displayed the temperature-sensitive phenotype.     

Elimination of Mange Mites <Emphasis Type="Italic">Sarcoptes scabiei</Emphasis> var. <Emphasis Type="Italic">suis</Emphasis>from Two Naturally Infested Danish Sow Herds Using a Single Injection Regime with Doramectin

Attempts to eliminate Sarcoptes scabiei var. suis were made in 2 naturally infested sow herds, by intramuscular (IM) injection of doramectin (Dectomax^®, Pfizer, New York, USA). A single injection strategy was used. In one of the herds, the environment was treated with an acaricide following dry cleaning of floors, walls and equipment. In the second herd, no environmental treatment was performed. Results were measured by skin lesion scoring, ear scrapings to show Sarcoptes scabiei var. suis, and calculating rubbing index throughout the observation period of 20 months following treatment. Skin lesion scores decreased and stayed low following treatment for the entire observation period. Live Sarcoptes scabiei var. suis mites were isolated prior to treatment from both herds, but not following treatment. Rubbing index decreased following treatment, but was occasionally at or above 0.4. The results of these studies indicate that elimination of Sarcoptes scabiei var. suis from 2 naturally infested herds was successful, using doramectin in a single injection strategy. Precautions must be taken to ensure adequate dosing of every pig, and to avoid reinfestation due to poor biosecurity.     

Photosynthetic electron transport adjustments in overwintering Scots pine (Pinus sylvestris L.)

As shown before [C. Ottander et al. (1995) Planta 197:176-183], there is a severe inhibition of the photosystem (PS) II photochemical efficiency of Scots pine (Pinus sylvestris L.) during the winter. In contrast, the in vivo PSI photochemistry is less inhibited during winter as shown by in vivo measurements of deltaA820/A820 (P700+). There was also an enhanced cyclic electron transfer around PSI in winter-stressed needles as indicated by 4-fold faster reduction kinetics of P700+. The differential functional stability of PSII and PSI was accompanied by a 3.7-fold higher intersystem electron pool size, and a 5-fold increase in the stromal electron pool available for P700+ reduction. There was also a strong reduction of the QB band in the thermoluminescence glow curve and markedly slower Q-A re-oxidation in needles of winter pine, indicating an inhibition of electron transfer between QA and QB. The data presented indicate that the plastoquinone pool is largely reduced in winter pine, and that this reduced state is likely to be of metabolic rather than photochemical origin. The retention of PSI photochemistry, and the suggested metabolic reduction of the plastoquinone pool in winter stressed needles of Scots pine are discussed in terms of the need for enhanced photoprotection of the needles during the winter and the role of metabolically supplied energy for the recovery of photosynthesis from winter stress in evergreens.     

Identification of rate-limiting steps in cephalosporin C biosynthesis in Cephalosporium acremonium: a theoretical analysis

Summary A kinetic model describing the biosynthesis of celphalosporin C in Cephalosporium acremonium has been developed to identify the rate-limiting step(s). Using this model and in-vitro kinetic data of the biosynthetic enzymes, the production kinetics of cephalosporin C were examined theoretically. The predicted time profile of the specific production rate during batch culture is in good agreement with that of experimental results published previously. Sensitivity analysis indicates that -(l--aminoadipyl)-l-cysteinyl-d-valine (ACV) synthetase is the rate-limiting enzyme. Our analysis also predicts that increasing ACV synthetase enhances the production rate initially until expandase/hydroxylase becomes rate-limiting. Furthermore, increasing expandase/hydroxylase reduces the accumulation of penicillin N, and thus, enhances the production of cephalosporin C. Based on our analysis, amplifying both ACV synthetase and expandase/hydroxylase concurrently should enhance the production rate to a great extent.Correspondence to: W. S. Hu